Expression Profiling Mark Voorhies 4/3/2012 Mark Voorhies - PowerPoint PPT Presentation

Expression Profiling Mark Voorhies 4/3/2012 Mark Voorhies Expression Profiling

It’s hard work at times, but you have to be realistic. If you have a large database with many variables and your goal is to get a good understanding of the interrelationships, then, unless you get lucky, this complex structure is bound to require some hard work to understand. Bill Cleveland and Rick Becker http://stat.bell-labs.com/project/trellis/interview.html Mark Voorhies Expression Profiling

Expression Profiling Why profile transcription ? Mark Voorhies Expression Profiling

Expression Profiling Why profile transcription ? Major mode of regulation Due to feedback, “shadows” other modes of regulation Thanks to Watson-Crick base pairing, we can assay arbitrary nucleic acids in a uniform way Mark Voorhies Expression Profiling

Expression Profiling Workflow Mark Voorhies Expression Profiling

Expression Profiling Analysis Mark Voorhies Expression Profiling

Sample Preparation Mark Voorhies Expression Profiling

Transforming Ratios Mark Voorhies Expression Profiling

Transforming Ratios Mark Voorhies Expression Profiling

Transforming Ratios M 1 = M 3 / M 2 Mark Voorhies Expression Profiling

Transforming Ratios log 2 M 1 = log 2 M 3 − log 2 M 2 Mark Voorhies Expression Profiling

The CDT file format Minimal CLUSTER input Cluster3 CDT output Tab delimited ( \ t) UNIX newlines ( \ n) Missing values → empty cells Mark Voorhies Expression Profiling

Comparing all measurements for two genes Comparing two expression profiles (r = 0.97) ● ● 5 ● ● ● YFG1 log2 relative expression ● ● ● ● ● ● ● ● ● ● ● 0 ● ● ● ● ● ● ● ● ● −5 ● ● ● ● ● −5 0 5 TLC1 log2 relative expression Mark Voorhies Expression Profiling

Comparing all genes for two measurements ● ● ● ● ● ● ● ● 5 ● ● ● ● ● ● ● ● Array 2, log2 relative expression ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● 0 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −5 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −10 ● ● −10 −5 0 5 10 Array 1, log2 relative expression Mark Voorhies Expression Profiling

Comparing all genes for two measurements Euclidean Distance ● ● ● ● ● ● ● ● 5 ● ● ● ● ● ● ● ● Array 2, log2 relative expression ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● 0 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −5 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −10 ● ● −10 −5 0 5 10 Array 1, log2 relative expression Mark Voorhies Expression Profiling

Comparing all genes for two measurements Uncentered Pearson ● ● ● ● ● ● ● ● 5 ● ● ● ● ● ● ● ● Array 2, log2 relative expression ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● 0 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −5 ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● ● −10 ● ● −10 −5 0 5 10 Array 1, log2 relative expression Mark Voorhies Expression Profiling

Measure all pairwise distances under distance metric Mark Voorhies Expression Profiling

Hierarchical Clustering Mark Voorhies Expression Profiling

Hierarchical Clustering Mark Voorhies Expression Profiling

Hierarchical Clustering Mark Voorhies Expression Profiling

Hierarchical Clustering Mark Voorhies Expression Profiling

Hierarchical Clustering Mark Voorhies Expression Profiling

Using the Cluster3 GUI Mark Voorhies Expression Profiling

Load your data Mark Voorhies Expression Profiling

Choose distance function Mark Voorhies Expression Profiling

Choose linking method Mark Voorhies Expression Profiling

Using JavaTreeView Mark Voorhies Expression Profiling

Adjust pixel settings for global view Mark Voorhies Expression Profiling

Adjust pixel settings for global view Mark Voorhies Expression Profiling

Select annotation columns Mark Voorhies Expression Profiling

Select annotation columns Mark Voorhies Expression Profiling

Select URL for gene annotations Mark Voorhies Expression Profiling

Select URL for gene annotations Mark Voorhies Expression Profiling

Activate and detach annotation window Mark Voorhies Expression Profiling

Activate and detach annotation window Mark Voorhies Expression Profiling

Activate and detach annotation window Mark Voorhies Expression Profiling

Homework Compare the effects of different distance metrics and clustering algorithms on the data from the Eisen paper (note that the GORDER column for the human data will make comparison easier). Practice annotating clusters in JavaTreeView. Try to find the annotated yeast clusters from the paper. Follow the links to SGD to see if the annotations for these genes have changed in the past decade. Read Bioinformatics 20:3710 Reminder: we are in HSW-532 tomorrow! Mark Voorhies Expression Profiling